This proposal describes a five year career development program whose goal is to prepare Dr. Julie Zikherman for a career as an independent investigator. This program will build on Dr. Zikherman's experience in clinical rheumatology, basic developmental biology and basic immunology in order to expand an emerging research program and provide her with further expertise in the design and conduct of cellular and molecular immunology research. The long-term goal is to establish an independent laboratory dedicated to the study of basic mechanisms of autoimmune disease pathogenesis, with a focus on how antigen receptor signaling can establish and subvert immune tolerance. The central guidance and training environment for this project will be provided by the mentor, Dr. Art Weiss, Engleman Distinguished Professor of Medicine, and Division Chief of Rheumatology at UCSF, as well as an expert in T cell receptor signaling. The training plan includes formal course work and didactic training through attendance of seminar series, journal clubs, UCSF retreats and scientific meetings, as well as guidance by an advisory committee of established scientists, and a research program that will provide additional technical training along with new directions for independent investigation. The studies proposed in this grant are intended to elucidate how altered T cell receptor signaling can subvert mechanisms of central tolerance and produce autoimmunity in humans. In her preliminary data, Dr. Zikherman has generated an allelic series of mice expressing distinct levels of CD45, and demonstrated that varying CD45 dose produces stable titration of TCR signaling intensity as well as quantitative changes in thymic output. In the research proposal, Dr. Zikherman will take advantage of this novel system to study how titration of TCR signaling intensity regulates thymic cell fate, transcriptional machinery and ultimately functional T cell repertoire. In the first part of the proposal, Dr. Zikherman will use genetic approaches and bone marrow chimeras to define the regulation of positive and negative selection in the allelic series. In the second part she will characterize the molecular phenotype of allelic series thymocytes by defining expression of key transcriptional regulators of positive selection. In the final part of the proposal, Dr. Zikherman will capitalize on the ability to track individual autoreactive cells in the context of the Aire-/- model of organ-specific autoimmunity to study how TCR signaling affects functional T cell repertoire. Together, these experiments will serve to characterize the cellular, molecular, and functional consequences of altered TCR signaling during T cell development. These studies will illuminate the mechanisms by which altered TCR signaling undermines central tolerance to produce disease. In addition to addressing the questions posed in this research proposal, the studies outlined above will serve as a foundation for future work that will enable Dr. Zikherman to become an independent investigator.